Psychiatr Q DOI 10.1007/s11126-013-9281-3 ORIGINAL PAPER
N-Acetylcysteine and Metabotropic Glutamate Receptors: Implications for the Treatment of Schizophrenia: A Literature review Adam Daniel Zavodnick • Rizwan Ali
Ó Springer Science+Business Media New York 2014
Abstract The objective of this study is to review the available data regarding metabotropic glutamate receptors in the pathology of Schizophrenia. Further, to investigate the potential utility of N-acetylcysteine as it relates to metabotropic glutamate receptors. A PubMed based literature review was conducted using keywords related to glutamate receptors, Schizophrenia and N-acetylcysteine from June 2012 through August of 2012. Relevant cited references of selected articles were also reviewed. The knowledge base regarding glutamate receptors, both ionotropic and metabotropic is rapidly expanding. New agonists of various subsets of metabotropic glutamate receptors are available and have demonstrated potential utility in animal models. N-acetylcysteine indirectly stimulates presynaptic metabotropic glutamate receptors and has shown efficacy in two double-blind randomized controlled trials. Metabotropic glutamate receptors contribute to an understanding of glutamate dysfunction in Schizophrenia. Agents which lead to stimulation of metabotropic glutamate receptors, inclusive of N-acetylcysteine show promise as novel agents in the treatment of this disorder. An understanding of the various metabotropic glutamate receptors will be a growing necessity as agents which target them continue to emerge and enter clinical trials. Keywords mglu
N-Acetylcysteine Metabotropic glutamate receptor Schizophrenia
A. D. Zavodnick (&) Psychiatry Intern, Department of Psychiatry and Behavioral Medicine, Carilion Clinic – Virginia Tech Carilion School of Medicine, Roanoke, VA, USA e-mail: [email protected] R. Ali Psychiatry Service, Veterans Affairs Medical Center, 1970 Roanoke Boulevard – 116A7, Salem, VA 24153, USA
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Introduction Disease models of Schizophrenia which emphasize dysfunction of the Glutamate system have been growing in popularity. These models were constructed in response to observations of cognitive deficits, hyper locomotion and stereotypies in animals and humans after receiving agents that interfered with the glutamatergic NMDA receptor (Fig. 1) [1–3]. In further support of this, blockade of the NMDA receptor has been shown to worsen both the positive and negative symptoms of psychosis in patients with Schizophrenia [4]. N-acetylcysteine is one of several agents currently available, which have effects on glutamatergic neurotransmission. It has been receiving increasing amounts of attention as it has shown utility in multiple neurological and psychiatric diseases inclusive of schizophrenia [5, 6]. This article will review glutamate mediated neurotransmission, glutamate receptor taxonomy and glutamatergic dysfunction as it relates to Schizophrenia (the NMDA receptor hypofunction hypothesis). It will conclude by exploring the potential utility of Nacetylcysteine in the treatment of Schizophrenia.
Methods A PubMed based literature review was conducted using keywords related to glutamate receptors, Schizophrenia and N-acetylcysteine in various combinations from June 2012 through August of 2012. Hundreds of articles were recovered and were reviewed based on their relevance to the objectives of this review article. Relevant cited references of selected articles were also reviewed for additional thoroughness.
Results Glutamate: An Overview Glutamate is thought to be the most abundant stimulatory neurotransmitter in the CNS, estimated to be present in up to 80 % of all synapses [7]. Importantly, the dynamics of glutamatergic neurotransmission are significantly different from serotonergic and dopaminergic neurotransmission. An introduction to glutamatergic neurotransmission with an emphasis on the diverse array of glutamate receptors will serve as a jumping off point from which to discuss the NMDA hypofunction disease model of Schizophrenia. Glutamate Receptors There are two main groups of Glutamate receptors in the human CNS. These include the ionotropic (iGlu) and metabotropic (mGlu) glutamate receptors [2]. Ionotropic receptors are so named because activation leads directly to the passage of ions through protein channels, resulting in a depolarization of the cell membrane. Ionotropic receptors are subdivided into the alpha-amino-3-hydroxy-5-methyl-4-isoxazole propionic acid (AMPA), Kainate (KA) and N-methyl-D-aspartic acid (NMDA) receptors, respectively. The individual receptors have been named for the compounds which were found to activate them, e.g. NMDA preferentially activates the NMDA receptor [2].
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Fig. 1 Pathways of metabotropic glutamate receptors
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AMPA and KA Receptors AMPA and KA receptors mediate fast EPSCs. As opposed to the NMDA receptors, they are able to function at resting cell polarity, i.e. they are not voltage dependent. Importantly, there is significant variability in the expression of AMPA receptors, due to heterogeneity in subunit composition as well as post transcriptional mRNA editing. Autoradiographic studies have shown AMPA receptors located diffusely throughout the CNS with high densities in the CA 1 straitum radiatum, dentate gyrus, superficial layers of the neocortex and the molecular layer of the cerebellum [2]. NMDA Receptors The NMDA receptor is functionally and compositionally distinct from the AMPA receptor. In addition to glutamate it also requires binding of glycine at a distinct modulatory site in order to be activated. It has other ancillary binding domains for zinc and polyamines, which inhibit and enhance receptor activity respectively. The glutamate binding site is not entirely specific as other amino acids such as L-homocysteic acid, L-aspartic acid and quinolinic acid can also agonize it. The NMDA receptor houses a magnesium ion which serves to block the channel during periods of resting cell polarity. It therefore requires cell depolarization from AMPA receptor activation in order to function, thus it is voltage dependent [2]. Given that it has higher affinity for glutamate than the AMPA receptor, it is able to be located more remotely than AMPA receptors on the postsynaptic membrane [7]. When activated, NMDA receptors allows the passage of sodium and calcium into the cell. Understimulation has been implicated in Schizophrenia while excitotoxicity secondary to excessive activation can lead to cell death and has been implicated in Alzheimer’s disease [2]. As with the AMPA receptor, there is configurational variability in the expression of the NMDA receptor, due in part to subunit composition and splice variation [2]. The NMDA receptor is also heavily influenced by the milieu in which it exists. For example, kynurenic acid (KYNA) a metabolite of tryptophan degradation has been shown to inhibit the NMDA receptor and has been demonstrated to be more prevalent in the prefrontal cortex of patients with Schizophrenia [8]. MK-801, Phencylidine and the dissociative anesthetic Ketamine exert their mechanisms of action by blocking the NMDA receptor and the deficits they produced helped in the develop the NMDA hypofunction disease model of Schizophrenia [1]. Finally, metabotropic glutamate receptors, which are reviewed below, influence the NMDA receptor as well. Given the ubiquity of the NMDA receptor, it is not surprising that dysfunction involving the receptor has been implicated in a number of distinct neurologic and psychiatric disorders inclusive of Schizophrenia [2]. In keeping with this, there are currently three NMDA receptor antagonists with FDA indications in distinct disorders—Riluzole for ALS, Acamprosate for alcohol dependence and Memantine for Alzheimer’s Disease [2, 9]. Metabotropic Glutamate Receptors Metabotropic glutamate receptors (mGluRs) are a distinct class of G-proteins with no known sequence homology to other GPCRs associated with other neurotransmitters. Instead, mGluRs are similar to the calcium sensor receptor of the parathyroid gland and the
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GABAb receptor. There are 8 known mGluRs (mGluR1—8), categorized into three divisions (type I, II and III) based on sequence homology, localization and second messenger utilization [1]. mGlu Type I Receptors mGluR type I receptors include mGluR1 and mGluR5, the activation of which leads to enhanced activity of the NMDA receptor. Agonism of the mGlu5 receptor has been demonstrated to ameliorate cognitive deficits induced by NMDA receptor blockade in an animal study [10]. mGluR Type II Receptors mGluR type II receptors, also known as mGlu2/3 receptors, include mGlu2 and mGlu3 receptors. When stimulated, they lead to inhibited production of cAMP and suppress cell depolarization, thus decreasing glutamate release from presynaptic nerve terminals. These two receptors differ from one another based on location within the CNS and in the synapse. While both can be located postsynaptically, radiotracer studies have found mGlu2 receptors to be often located presynaptically, functioning as an autoreceptor with wide distribution within the cortical-striato-thalamic re-entrant loops and limbic related circuitry [11]. In the prefrontal cortex, mGlu2 receptors are found principally in layers I and Va, overlapping with 5HT2A receptor distribution. In keeping with this, mGlu2 stimulation has been shown to attenuate behavioral abnormalities induced by hallucinogens as well of NMDA receptor antagonists. Examples of synthetic agents which agonize mGlu2 receptors include LY354740, LY404039, LY379268, MGS0008 and MGS0028 [12]. mGlu3 receptors are located post synaptically and are found as heteroreceptors on glia as well as on GABAergic interneurons. Distribution within the CNS is more diffuse than for the mGlu2 receptor and decreased expression has been noted in post mortem findings of patients with Schizophrenia. Despite this, it appears that mGlu2 agonism may be more beneficial in the treatment of psychosis than mGlu3 agonism [11, 12]. Type III mGlu Receptors Type III mGlu receptors include mGluR4, 6, 7 and 8. They comprise the largest group of mGlu receptors and have been less thoroughly investigated. mGluR6 is postsynaptic, restricted to the retina and mediates synaptic transmission at retinal ON bipolar cells. The remaining receptors mGlu 4, 7 and 8 are presynaptic, widely distributed in the CNS and function as inhibitory autoreceptors on both glutamatergic and GABAergic neurons. Recent investigation into the role of mGlu7 through the use of the modulator N,N0 -dibenzhydrylethane-1,2-diamine dihydrochloride (AMN082) suggests potential utility of group III agonists in psychiatric conditions inclusive of anxiety [13]. Glia Glial cells are actively involved in the regulation of glutamatergic neurotransmission through multiple mechanisms. They clear glutamate released from the synapse, transforming it to glutamine before releasing it back to the presynaptic glutamate neurons. In addition, it regulates glutamatergic tone via the cystine-glutamate antiporter, a protein
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which is able to exchange glutamate and cystine in a 1:1 stoichiometric ratio. This protein thus enables glia to release glutamate extracellularly, where it can stimulate presynaptic mGlu2 receptors. As will be discussed below, the pharmacologic augmentation of this antiporter is a putative mechanism of action for n-acetyl-cysteine, the administration of which leads to extrasynaptic glutamate release and subsequent presynaptic autoreceptor stimulation [7, 14]. NMDA Receptor Hypofunction as a Disease Model of Schizophrenia As mentioned above, abnormal NMDA receptor stimulation has been implicated in a number of neuronal diseases. However, NMDA receptor hypofunction rather than hyperfunction has been implicated in the pathogenesis of Schizophrenia. This NMDA receptor hypofunction model of Schizophrenia followed the findings of abnormal glutamate concentrations in certain postmortem studies of patients with Schizophrenia and induction of cognitive dysfunction following administration of NMDA receptor antagonists such as MK 801, Phencyclidine and Ketamine. In keeping with this were the findings that compounds that augment the function of the NMDA receptor (D-serine, cycloserine) ameliorate the cognitive dysfunction seen in Schizophrenia [13]. Clozapine, an agent uniquely beneficial in the treatment of refractory Schizophrenia, has also been implicated in facilitation of NMDA receptor activation [2]. As NMDA receptor mediated neurotransmission is complex, there are a number of potential vulnerabilities to disturbance. These include interference from exogenous substances, altered composition of the receptor or abnormal interactions with mGlu receptors. Consistent with this, abnormal expression of mGlu2/3 receptors as well as polymorphisms in the gene which encodes mGlu3 have been noted in post mortem analysis of patients with Schizophrenia [13]. As NMDA receptors are found postsynaptically on cells which suppress presynaptic glutamate release (GABAergic interneurons, regulatory glial cells), underfunction of the NMDA receptor, regardless of etiology would be expected to result in an increase in glutamate release at the synapse because of an interrupted negative feedback loop [8]. This could potentially lead to an abnormally increased ratio of post synaptic AMPA mediated effects relative to NMDA mediated effects. Abnormal AMPA/NMDA stimulation ratio has been shown to lead to abnormalities in a computer generated model of neuron firing [15]. One strategy for rectification of this hyperglutamatergic tone would be through stimulation of metabotropic glutamate receptors. Stimulation of group I metabotropic glutamate receptors would facilitate NMDA function while stimulation of group II and group III metabotropic glutamate receptors should lead directly to suppression of presynaptic glutamate release. Experiments using agents which act as mGlu receptor agonists lend support for this hypothesis [10, 12, 13]. Alternatively, N-acetylcysteine indirectly leads to the stimulation of group II metabotropic receptors via the glial cystine/glutamate antiporter discussed below [7]. N-Acetylcysteine Long used as a mucolytic and in the treatment of acetaminophen overdose [16], N-acetylcysteine has been investigated in an incredibly broad array of disorders over the years from alcoholic hepatitis [17] to iodine-induced nephropathy [18]. Its mechanism of action in detoxification involves the generation of glutathione (GSH), a potent antioxidant in the mammalian body. As abnormalities in inflammation in general and glutathione specifically
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have been implicated in Schizophrenia [19, 20], this may be a distinct mechanism of action potentially unrelated to metabotropic glutamate receptor stimulation. In support of a glutathione-deficient hypothesis of Schizophrenia, abnormal levels of glutathione and abnormalities in enzymes involved in glutathione synthesis have been noted in patients with Schizophrenia. Neurotransmitter abnormalities similar to those noted in Schizophrenia have been shown in glutathione deficient mice and normalization of these neurotransmitters have been shown with N-acetylcysteine supplementation [21]. As it relates to glutamatergic neurotransmission, N-acetylcysteine is believed to promote the stimulation of group II metabotropic autoreceptors through the extracellular generation of cystine, which subsequently leads to increased glutamate release from astrocytes via the Cystine-Glutamate antiporter [7, 14]. N-acetylcysteine has been shown to ameliorate social and cognitive deficits in PCP treated rodents, while blockade of mGlu2 receptors prevents this ameliorative effect [21]. The current body of evidence in support of n-acetyl-cysteine for Schizophrenia include animal studies [21–23], a case report [24] and two double blind studies [25, 26]. The latter two are discussed below. Clinical Studies in Patients with Schizophrenia The first published double blind randomized control trial investigating N-acetylcysteine was published in 2007 by Lavoie et al. [25]. It was small, with only seven patients completing the study. The primary outcome was change in mismatch negativity (MMN), a measure of cognitive deficit in Schizophrenia thought to be associated with impaired NMDA receptor function. Patients were compared to age and sex matched healthy controls at baseline and then were randomized to receive either placebo or NAC 1 gram twice a day for 60 days and then switched to the opposite condition. The authors demonstrated a statistically significant improvement compared to placebo in MMN, although there were no significant changes in global level of functioning. The second double blind study was published by Berk et al. in 2008, in which 84 patients with Schizophrenia were followed for 4 weeks and treated with either N-acetylcysteine 1 g twice a day or placebo for 24 weeks. This study showed improvement in global scales, inclusive of PANSS total, PANSS negative, PANSS general, CGI-Severity and CGI-Improvement but not PANSS positive. Akathisia was also improved in treated subjects. However, not all studies have been positive, Gunduz demonstrated that N-acetylcysteine pretreatment increased rather than decreased MMN in healthy volunteers treated with Ketamine, although it did also increase target and novelty P3 amplitudes, suggesting potential efficacy in certain domains [27]. Encouraging findings in other potentially related psychiatric conditions such as bipolar disorder have also been accumulating [28].
Conclusion The ubiquity of the glutamate system in the mammalian CNS is matched by its complexity. Abnormalities in various subtypes of glutamate receptors have been implicated in a wide variety of neuropsychiatric disease states inclusive of Schizophrenia. N-acetylcysteine is one of a growing number of agents which influence the metabotropic glutamate receptors. With encouraging preliminary evidence, future research is warranted.
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References 1. Moghaddam B, Javitt D: From revolution to evolution: The glutamate hypothesis of schizophrenia and its implication for treatment. Neuropsychopharmacology 37(1):4–15, 2012. 2. Coyle J, Leski M, Morrison J, et al.: The diverse roles of L-glutamic acid in brain signal transduction. In: Davis K, Charney D, Coyle J, Nemeroff C (Eds) Neuropsychopharmacology (The Fifth Generation of Progress). Philadelphia: Lippincott, Williams & Wilkins, pp. 71–90, 2002. 3. Adell A, Jime´nez-Sa´nchez L, Lo´pez-Gil X, Romo´n T: Is the acute NMDA receptor hypofunction a valid model of schizophrenia? Schizophrenia Bulletin 38(1):9–14, 2012. 4. Goff DC, Coyle JT: The emerging role of glutamate in the pathophysiology and treatment of schizophrenia. American Journal of Psychiatry 158(9):1367–1377, 2001. 5. Sansone RA, Sansone LA: Getting a knack for NAC: N-acetyl-cysteine. Innovations in Clinical Neuroscience 8(1):10–14, 2011. 6. Dean O, Giorlando F, Berk M: N-acetylcysteine in psychiatry: Current therapeutic evidence and potential mechanisms of action. Journal of Psychiatry & Neuroscience 36(2):78–86, 2011. 7. Bridges R, Lutgen V, Lobner D, Baker DA: Thinking outside the cleft to understand synaptic activity: Contribution of the cystine-glutamate antiporter (System xc-) to normal and pathological glutamatergic signaling. Pharmacological Reviews 64(3):780–802, 2012. 8. Sathyasaikumar KV, Stachowski EK, Wonodi I, Roberts RC, Rassoulpour A, McMahon RP, Schwarcz R: Impaired kynurenine pathway metabolism in the prefrontal cortex of individuals with schizophrenia. Schizophrenia Bulletin 37(6):1147–1156, 2011. 9. Olive MF, Cleva RM, Kalivas PW, Malcolm RJ: Glutamatergic medications for the treatment of drug and behavioral addictions. Pharmacology Biochemistry and Behavior 100(4):801–810, 2012. 10. Stefani MR, Moghaddam B: Activation of type 5 metabotropic glutamate receptors attenuates deficits in cognitive flexibility induced by NMDA receptor blockade. European Journal of Pharmacology 639(1–3):26–32, 2010. 11. Marek GJ: Metabotropic glutamate2/3 (mGlu2/3) receptors, schizophrenia and cognition. European Journal of Pharmacology 639(1–3):81–90, 2010. 12. Chaki S: Group II metabotropic glutamate receptor agonists as a potential drug for schizophrenia. European Journal of Pharmacology 639(1–3):59–66, 2010. 13. O’Connor RM, Finger BC, Flor PJ, Cryan JF: Metabotropic glutamate receptor 7: At the interface of cognition and emotion. European Journal of Pharmacology 639(1–3):123–131, 2010. 14. Moran MM, McFarland K, Melendez RI, Kalivas PW, Seamans JK: Cystine/glutamate exchange regulates metabotropic glutamate receptor presynaptic inhibition of excitatory transmission and vulnerability to cocaine seeking. The Journal of Neuroscience 25(27):6389–6393. 15. Wolf JA, Moyer JT, Lazarewicz MT, Contreras D, Benoit-Marand M, O’Donnell P, Finkel LH: NMDA/ AMPA ratio impacts state transitions and entrainment to oscillations in a computational model of the nucleus accumbens medium spiny projection neuron. The Journal of Neuroscience 25(40):9080–9095, 2005. 16. Smilkstein MJ, Knapp GL, Kulig KW, Rumack BH: Efficacy of oral N-acetylcysteine in the treatment of acetaminophen overdose. Analysis of the national multicenter study (1976 to 1985). The New England Journal of Medicine 319(24):1557–1562, 1988. 17. Nguyen-Khac E, Thevenot T, Piquet MA, Benferhat S, Goria O, Chatelain D, Tramier B, Dewaele F, Ghrib S, Rudler M, Carbonell N, Tossou H, Bental A, Bernard-Chabert B, Dupas JL; AAH-NAC Study Group: Glucocorticoids plus N-acetylcysteine in severe alcoholic hepatitis. The New England Journal of Medicine 365(19):1781–1789, 2011. 18. Marenzi G, Assanelli E, Marana I, Lauri G, Campodonico J, Grazi M, De Metrio M, Galli S, Fabbiocchi F, Montorsi P, Veglia F, Bartorelli AL: N-acetylcysteine and contrast-induced nephropathy in primary angioplasty. The New England Journal of Medicine 354(26):2773–2782, 2006. 19. Bosˇkovic´ M, Vovk T, Kores Plesnicˇar B, Grabnar I: Oxidative stress in schizophrenia. Current Neuropharmacology 9(2):301–312, 2011. 20. Gysin R, Kraftsik R, Sandell J, Bovet P, Chappuis C, Conus P, Deppen P, Preisig M, Ruiz V, Steullet P, Tosic M, Werge T, Cue´nod M, Do KQ: Impaired glutathione synthesis in schizophrenia: Convergent genetic and functional evidence. Proceedings of the National Academy of Sciences of the United States of America 104(42):16621–16666, 2007. 21. Shungu DC: N-acetylcysteine for the treatment of glutathione deficiency and oxidative stress in schizophrenia. Biological Psychiatry 71(11):937–938, 2012.
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Adam Daniel Zavodnick MD is a psychiatry resident with the Department of Psychiatry and Behavioral Medicine Carilion Clinic—Virginia Tech Carilion School of Medicine, Roanoke, Virginia Rizwan Ali MD is a staff psychiatrist at Veterans Affairs Medical Center, Salem, Virginia and Assistant Professor with the Department of Psychiatry and Behavioral Medicine Carilion Clinic—Virginia Tech Carilion School of Medicine, Roanoke, Virginia
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N-Acetylcysteine and Metabotropic Glutamate Receptors: Implications for the Treatment of Schizophrenia: A Literature review Adam Daniel Zavodnick • Rizwan Ali
Ó Springer Science+Business Media New York 2014
Abstract The objective of this study is to review the available data regarding metabotropic glutamate receptors in the pathology of Schizophrenia. Further, to investigate the potential utility of N-acetylcysteine as it relates to metabotropic glutamate receptors. A PubMed based literature review was conducted using keywords related to glutamate receptors, Schizophrenia and N-acetylcysteine from June 2012 through August of 2012. Relevant cited references of selected articles were also reviewed. The knowledge base regarding glutamate receptors, both ionotropic and metabotropic is rapidly expanding. New agonists of various subsets of metabotropic glutamate receptors are available and have demonstrated potential utility in animal models. N-acetylcysteine indirectly stimulates presynaptic metabotropic glutamate receptors and has shown efficacy in two double-blind randomized controlled trials. Metabotropic glutamate receptors contribute to an understanding of glutamate dysfunction in Schizophrenia. Agents which lead to stimulation of metabotropic glutamate receptors, inclusive of N-acetylcysteine show promise as novel agents in the treatment of this disorder. An understanding of the various metabotropic glutamate receptors will be a growing necessity as agents which target them continue to emerge and enter clinical trials. Keywords mglu
N-Acetylcysteine Metabotropic glutamate receptor Schizophrenia
A. D. Zavodnick (&) Psychiatry Intern, Department of Psychiatry and Behavioral Medicine, Carilion Clinic – Virginia Tech Carilion School of Medicine, Roanoke, VA, USA e-mail: [email protected] R. Ali Psychiatry Service, Veterans Affairs Medical Center, 1970 Roanoke Boulevard – 116A7, Salem, VA 24153, USA
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Introduction Disease models of Schizophrenia which emphasize dysfunction of the Glutamate system have been growing in popularity. These models were constructed in response to observations of cognitive deficits, hyper locomotion and stereotypies in animals and humans after receiving agents that interfered with the glutamatergic NMDA receptor (Fig. 1) [1–3]. In further support of this, blockade of the NMDA receptor has been shown to worsen both the positive and negative symptoms of psychosis in patients with Schizophrenia [4]. N-acetylcysteine is one of several agents currently available, which have effects on glutamatergic neurotransmission. It has been receiving increasing amounts of attention as it has shown utility in multiple neurological and psychiatric diseases inclusive of schizophrenia [5, 6]. This article will review glutamate mediated neurotransmission, glutamate receptor taxonomy and glutamatergic dysfunction as it relates to Schizophrenia (the NMDA receptor hypofunction hypothesis). It will conclude by exploring the potential utility of Nacetylcysteine in the treatment of Schizophrenia.
Methods A PubMed based literature review was conducted using keywords related to glutamate receptors, Schizophrenia and N-acetylcysteine in various combinations from June 2012 through August of 2012. Hundreds of articles were recovered and were reviewed based on their relevance to the objectives of this review article. Relevant cited references of selected articles were also reviewed for additional thoroughness.
Results Glutamate: An Overview Glutamate is thought to be the most abundant stimulatory neurotransmitter in the CNS, estimated to be present in up to 80 % of all synapses [7]. Importantly, the dynamics of glutamatergic neurotransmission are significantly different from serotonergic and dopaminergic neurotransmission. An introduction to glutamatergic neurotransmission with an emphasis on the diverse array of glutamate receptors will serve as a jumping off point from which to discuss the NMDA hypofunction disease model of Schizophrenia. Glutamate Receptors There are two main groups of Glutamate receptors in the human CNS. These include the ionotropic (iGlu) and metabotropic (mGlu) glutamate receptors [2]. Ionotropic receptors are so named because activation leads directly to the passage of ions through protein channels, resulting in a depolarization of the cell membrane. Ionotropic receptors are subdivided into the alpha-amino-3-hydroxy-5-methyl-4-isoxazole propionic acid (AMPA), Kainate (KA) and N-methyl-D-aspartic acid (NMDA) receptors, respectively. The individual receptors have been named for the compounds which were found to activate them, e.g. NMDA preferentially activates the NMDA receptor [2].
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Fig. 1 Pathways of metabotropic glutamate receptors
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AMPA and KA Receptors AMPA and KA receptors mediate fast EPSCs. As opposed to the NMDA receptors, they are able to function at resting cell polarity, i.e. they are not voltage dependent. Importantly, there is significant variability in the expression of AMPA receptors, due to heterogeneity in subunit composition as well as post transcriptional mRNA editing. Autoradiographic studies have shown AMPA receptors located diffusely throughout the CNS with high densities in the CA 1 straitum radiatum, dentate gyrus, superficial layers of the neocortex and the molecular layer of the cerebellum [2]. NMDA Receptors The NMDA receptor is functionally and compositionally distinct from the AMPA receptor. In addition to glutamate it also requires binding of glycine at a distinct modulatory site in order to be activated. It has other ancillary binding domains for zinc and polyamines, which inhibit and enhance receptor activity respectively. The glutamate binding site is not entirely specific as other amino acids such as L-homocysteic acid, L-aspartic acid and quinolinic acid can also agonize it. The NMDA receptor houses a magnesium ion which serves to block the channel during periods of resting cell polarity. It therefore requires cell depolarization from AMPA receptor activation in order to function, thus it is voltage dependent [2]. Given that it has higher affinity for glutamate than the AMPA receptor, it is able to be located more remotely than AMPA receptors on the postsynaptic membrane [7]. When activated, NMDA receptors allows the passage of sodium and calcium into the cell. Understimulation has been implicated in Schizophrenia while excitotoxicity secondary to excessive activation can lead to cell death and has been implicated in Alzheimer’s disease [2]. As with the AMPA receptor, there is configurational variability in the expression of the NMDA receptor, due in part to subunit composition and splice variation [2]. The NMDA receptor is also heavily influenced by the milieu in which it exists. For example, kynurenic acid (KYNA) a metabolite of tryptophan degradation has been shown to inhibit the NMDA receptor and has been demonstrated to be more prevalent in the prefrontal cortex of patients with Schizophrenia [8]. MK-801, Phencylidine and the dissociative anesthetic Ketamine exert their mechanisms of action by blocking the NMDA receptor and the deficits they produced helped in the develop the NMDA hypofunction disease model of Schizophrenia [1]. Finally, metabotropic glutamate receptors, which are reviewed below, influence the NMDA receptor as well. Given the ubiquity of the NMDA receptor, it is not surprising that dysfunction involving the receptor has been implicated in a number of distinct neurologic and psychiatric disorders inclusive of Schizophrenia [2]. In keeping with this, there are currently three NMDA receptor antagonists with FDA indications in distinct disorders—Riluzole for ALS, Acamprosate for alcohol dependence and Memantine for Alzheimer’s Disease [2, 9]. Metabotropic Glutamate Receptors Metabotropic glutamate receptors (mGluRs) are a distinct class of G-proteins with no known sequence homology to other GPCRs associated with other neurotransmitters. Instead, mGluRs are similar to the calcium sensor receptor of the parathyroid gland and the
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GABAb receptor. There are 8 known mGluRs (mGluR1—8), categorized into three divisions (type I, II and III) based on sequence homology, localization and second messenger utilization [1]. mGlu Type I Receptors mGluR type I receptors include mGluR1 and mGluR5, the activation of which leads to enhanced activity of the NMDA receptor. Agonism of the mGlu5 receptor has been demonstrated to ameliorate cognitive deficits induced by NMDA receptor blockade in an animal study [10]. mGluR Type II Receptors mGluR type II receptors, also known as mGlu2/3 receptors, include mGlu2 and mGlu3 receptors. When stimulated, they lead to inhibited production of cAMP and suppress cell depolarization, thus decreasing glutamate release from presynaptic nerve terminals. These two receptors differ from one another based on location within the CNS and in the synapse. While both can be located postsynaptically, radiotracer studies have found mGlu2 receptors to be often located presynaptically, functioning as an autoreceptor with wide distribution within the cortical-striato-thalamic re-entrant loops and limbic related circuitry [11]. In the prefrontal cortex, mGlu2 receptors are found principally in layers I and Va, overlapping with 5HT2A receptor distribution. In keeping with this, mGlu2 stimulation has been shown to attenuate behavioral abnormalities induced by hallucinogens as well of NMDA receptor antagonists. Examples of synthetic agents which agonize mGlu2 receptors include LY354740, LY404039, LY379268, MGS0008 and MGS0028 [12]. mGlu3 receptors are located post synaptically and are found as heteroreceptors on glia as well as on GABAergic interneurons. Distribution within the CNS is more diffuse than for the mGlu2 receptor and decreased expression has been noted in post mortem findings of patients with Schizophrenia. Despite this, it appears that mGlu2 agonism may be more beneficial in the treatment of psychosis than mGlu3 agonism [11, 12]. Type III mGlu Receptors Type III mGlu receptors include mGluR4, 6, 7 and 8. They comprise the largest group of mGlu receptors and have been less thoroughly investigated. mGluR6 is postsynaptic, restricted to the retina and mediates synaptic transmission at retinal ON bipolar cells. The remaining receptors mGlu 4, 7 and 8 are presynaptic, widely distributed in the CNS and function as inhibitory autoreceptors on both glutamatergic and GABAergic neurons. Recent investigation into the role of mGlu7 through the use of the modulator N,N0 -dibenzhydrylethane-1,2-diamine dihydrochloride (AMN082) suggests potential utility of group III agonists in psychiatric conditions inclusive of anxiety [13]. Glia Glial cells are actively involved in the regulation of glutamatergic neurotransmission through multiple mechanisms. They clear glutamate released from the synapse, transforming it to glutamine before releasing it back to the presynaptic glutamate neurons. In addition, it regulates glutamatergic tone via the cystine-glutamate antiporter, a protein
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which is able to exchange glutamate and cystine in a 1:1 stoichiometric ratio. This protein thus enables glia to release glutamate extracellularly, where it can stimulate presynaptic mGlu2 receptors. As will be discussed below, the pharmacologic augmentation of this antiporter is a putative mechanism of action for n-acetyl-cysteine, the administration of which leads to extrasynaptic glutamate release and subsequent presynaptic autoreceptor stimulation [7, 14]. NMDA Receptor Hypofunction as a Disease Model of Schizophrenia As mentioned above, abnormal NMDA receptor stimulation has been implicated in a number of neuronal diseases. However, NMDA receptor hypofunction rather than hyperfunction has been implicated in the pathogenesis of Schizophrenia. This NMDA receptor hypofunction model of Schizophrenia followed the findings of abnormal glutamate concentrations in certain postmortem studies of patients with Schizophrenia and induction of cognitive dysfunction following administration of NMDA receptor antagonists such as MK 801, Phencyclidine and Ketamine. In keeping with this were the findings that compounds that augment the function of the NMDA receptor (D-serine, cycloserine) ameliorate the cognitive dysfunction seen in Schizophrenia [13]. Clozapine, an agent uniquely beneficial in the treatment of refractory Schizophrenia, has also been implicated in facilitation of NMDA receptor activation [2]. As NMDA receptor mediated neurotransmission is complex, there are a number of potential vulnerabilities to disturbance. These include interference from exogenous substances, altered composition of the receptor or abnormal interactions with mGlu receptors. Consistent with this, abnormal expression of mGlu2/3 receptors as well as polymorphisms in the gene which encodes mGlu3 have been noted in post mortem analysis of patients with Schizophrenia [13]. As NMDA receptors are found postsynaptically on cells which suppress presynaptic glutamate release (GABAergic interneurons, regulatory glial cells), underfunction of the NMDA receptor, regardless of etiology would be expected to result in an increase in glutamate release at the synapse because of an interrupted negative feedback loop [8]. This could potentially lead to an abnormally increased ratio of post synaptic AMPA mediated effects relative to NMDA mediated effects. Abnormal AMPA/NMDA stimulation ratio has been shown to lead to abnormalities in a computer generated model of neuron firing [15]. One strategy for rectification of this hyperglutamatergic tone would be through stimulation of metabotropic glutamate receptors. Stimulation of group I metabotropic glutamate receptors would facilitate NMDA function while stimulation of group II and group III metabotropic glutamate receptors should lead directly to suppression of presynaptic glutamate release. Experiments using agents which act as mGlu receptor agonists lend support for this hypothesis [10, 12, 13]. Alternatively, N-acetylcysteine indirectly leads to the stimulation of group II metabotropic receptors via the glial cystine/glutamate antiporter discussed below [7]. N-Acetylcysteine Long used as a mucolytic and in the treatment of acetaminophen overdose [16], N-acetylcysteine has been investigated in an incredibly broad array of disorders over the years from alcoholic hepatitis [17] to iodine-induced nephropathy [18]. Its mechanism of action in detoxification involves the generation of glutathione (GSH), a potent antioxidant in the mammalian body. As abnormalities in inflammation in general and glutathione specifically
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have been implicated in Schizophrenia [19, 20], this may be a distinct mechanism of action potentially unrelated to metabotropic glutamate receptor stimulation. In support of a glutathione-deficient hypothesis of Schizophrenia, abnormal levels of glutathione and abnormalities in enzymes involved in glutathione synthesis have been noted in patients with Schizophrenia. Neurotransmitter abnormalities similar to those noted in Schizophrenia have been shown in glutathione deficient mice and normalization of these neurotransmitters have been shown with N-acetylcysteine supplementation [21]. As it relates to glutamatergic neurotransmission, N-acetylcysteine is believed to promote the stimulation of group II metabotropic autoreceptors through the extracellular generation of cystine, which subsequently leads to increased glutamate release from astrocytes via the Cystine-Glutamate antiporter [7, 14]. N-acetylcysteine has been shown to ameliorate social and cognitive deficits in PCP treated rodents, while blockade of mGlu2 receptors prevents this ameliorative effect [21]. The current body of evidence in support of n-acetyl-cysteine for Schizophrenia include animal studies [21–23], a case report [24] and two double blind studies [25, 26]. The latter two are discussed below. Clinical Studies in Patients with Schizophrenia The first published double blind randomized control trial investigating N-acetylcysteine was published in 2007 by Lavoie et al. [25]. It was small, with only seven patients completing the study. The primary outcome was change in mismatch negativity (MMN), a measure of cognitive deficit in Schizophrenia thought to be associated with impaired NMDA receptor function. Patients were compared to age and sex matched healthy controls at baseline and then were randomized to receive either placebo or NAC 1 gram twice a day for 60 days and then switched to the opposite condition. The authors demonstrated a statistically significant improvement compared to placebo in MMN, although there were no significant changes in global level of functioning. The second double blind study was published by Berk et al. in 2008, in which 84 patients with Schizophrenia were followed for 4 weeks and treated with either N-acetylcysteine 1 g twice a day or placebo for 24 weeks. This study showed improvement in global scales, inclusive of PANSS total, PANSS negative, PANSS general, CGI-Severity and CGI-Improvement but not PANSS positive. Akathisia was also improved in treated subjects. However, not all studies have been positive, Gunduz demonstrated that N-acetylcysteine pretreatment increased rather than decreased MMN in healthy volunteers treated with Ketamine, although it did also increase target and novelty P3 amplitudes, suggesting potential efficacy in certain domains [27]. Encouraging findings in other potentially related psychiatric conditions such as bipolar disorder have also been accumulating [28].
Conclusion The ubiquity of the glutamate system in the mammalian CNS is matched by its complexity. Abnormalities in various subtypes of glutamate receptors have been implicated in a wide variety of neuropsychiatric disease states inclusive of Schizophrenia. N-acetylcysteine is one of a growing number of agents which influence the metabotropic glutamate receptors. With encouraging preliminary evidence, future research is warranted.
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Adam Daniel Zavodnick MD is a psychiatry resident with the Department of Psychiatry and Behavioral Medicine Carilion Clinic—Virginia Tech Carilion School of Medicine, Roanoke, Virginia Rizwan Ali MD is a staff psychiatrist at Veterans Affairs Medical Center, Salem, Virginia and Assistant Professor with the Department of Psychiatry and Behavioral Medicine Carilion Clinic—Virginia Tech Carilion School of Medicine, Roanoke, Virginia
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